While nonsense-mediated RNA decay (NMD) is an established mechanism to rapidly degrade select transcripts, the physiological regulation and biological significance of NMD are not well characterized. We previously demonstrated that NMD is inhibited in hypoxic cells. Here we show that the phosphorylation of the ␣ subunit of eukaryotic initiation factor 2 (eIF2␣) translation initiation factor by a variety of cellular stresses leads to the inhibition of NMD and that eIF2␣ phosphorylation and NMD inhibition occur in tumors. To explore the significance of this NMD regulation, we used an unbiased approach to identify approximately 750 NMD-targeted mRNAs and found that these mRNAs are overrepresented in stress response and tumorpromoting pathways. Consistent with these findings, the inhibition of NMD promotes cellular resistance to endoplasmic reticulum stress and encourages tumor formation. The transcriptional and translational regulations of gene expression by the microenvironment are established mechanisms by which tumor cells adapt to stress. These data indicate that NMD inhibition by the tumor microenvironment is also an important mechanism to dynamically regulate genes critical for the response to cellular stress and tumorigenesis.During tumorigenesis, a disorganized vasculature leads to amino acid and glucose deprivation, cellular hypoxia, the accumulation of reactive oxygen species (ROS), and various other stresses (5, 12). Cellular adaptation to the hostile tumor microenvironment requires the regulation of stress-induced genes (reviewed in reference 16). For example, the transcription factor ATF-4, upregulated in human tumors due to the stress-induced phosphorylation of the ␣ subunit of eukaryotic translation initiation factor 2 (eIF2␣), transactivates genes involved in amino acid metabolism, angiogenesis, and ROS attenuation (2,3,33). Cells that cannot phosphorylate eIF2␣ or that are deficient in ATF-4 and other stress-induced transcription factors do not form tumors in vivo (2, 13, 40), and therefore, a major goal in cancer biology has been to better understand and potentially target these adaptive mechanisms. However, while the translational and transcriptional responses that promote adaptation to the tumor microenvironment are well established, the role of mRNA stabilization in the cellular stress response has not been as thoroughly studied.Nonsense-mediated RNA decay (NMD) degrades up to 30% of all mutated protein-coding mRNAs, including those responsible for many genetic disorders, such as thalassemia, cystic fibrosis, and muscular dystrophy (11). During the processing of mammalian pre-mRNA, introns are excised and marked by an exon junction complex, which contains core NMD components. Newly synthesized mRNAs are thought to undergo a pioneering round of translation by a complex that includes eIF2␣ (6). When this translation complex pauses at a premature termination codon (PTC) upstream of an exon junction complex, the RNA helicase UPF1/Rent1, an essential component of the NMD process, is recruited and the...

Finding new therapies to assist in the treatment of cancer is a major challenge of clinical research. Small molecules that inhibit different molecular targets at the different levels of the MAPK pathway have been developed. Several MEK inhibitors have been examined in early-phase clinical trials and the current state of clinical results using these therapies is presented here.

The generation of beta-thalassemia (β-Thal) patient-specific induced pluripotent stem cells (iPSCs), subsequent homologous recombination-based gene correction of disease-causing mutations/deletions in the β-globin gene (HBB), and their derived hematopoietic stem cell (HSC) transplantation offers an ideal therapeutic solution for treating this disease. However, the hematopoietic differentiation efficiency of gene-corrected β-Thal iPSCs has not been well evaluated in the previous studies. In this study, we used the latest gene-editing tool, clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9), to correct β-Thal iPSCs; gene-corrected cells exhibit normal karyotypes and full pluripotency as human embryonic stem cells (hESCs) showed no off-targeting effects. Then, we evaluated the differentiation efficiency of the gene-corrected β-Thal iPSCs. We found that during hematopoietic differentiation, gene-corrected β-Thal iPSCs showed an increased embryoid body ratio and various hematopoietic progenitor cell percentages. More importantly, the gene-corrected β-Thal iPSC lines restored HBB expression and reduced reactive oxygen species production compared with the uncorrected group. Our study suggested that hematopoietic differentiation efficiency of β-Thal iPSCs was greatly improved once corrected by the CRISPR/Cas9 system, and the information gained from our study would greatly promote the clinical application of β-Thal iPSC-derived HSCs in transplantation.

Many of the gene mutations found in genetic disorders, including cancer, result in premature termination codons (PTCs) and the rapid degradation of their mRNAs by nonsense mediated RNA decay (NMD). We used virtual library screening (VLS) targeting a pocket in the SMG7 protein, a key component of the NMD mechanism, to identify compounds that disrupt the SMG7-UPF1 complex and inhibit NMD. Several of these compounds upregulated NMD targeted mRNAs at nanomolar concentrations with minimal toxicity in cell based assays. As expected, pharmacological NMD inhibition disrupted SMG7-UPF1 interactions. When used in cells with PTC mutated p53, pharmacological NMD inhibition combined with a PTC “read-through” drug led to restoration of full-length p53 protein, upregulation of p53 downstream transcripts, and cell death. These studies serve as proof-of-concept that pharmacological NMD inhibitors can restore mRNA integrity in the presence of PTC and be used as part of a strategy to restore full length protein in a variety of genetic diseases.